Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Identification of type A spermatogonia specific transcriptome in rainbow trout immature testis

ABSTRACT: To identify the transcripts preferentially expressed in type A spermatogonia in rainbow trout immature testis, we compared the transcriptome between type A spermatogonia and testicular somatic cells by microarry analysis. We used fluorescence-activated cell sorting (FACS) to isolate type A spermatogonia or testicular somatic cells from the fishes carrying the transgene, pvasa-GFP, which expresses GFP in spermatogonia. RNA from type A spermatogonia and from testicular somatic cells were hybridized to a microarray after Cy3 labeling.

ORGANISM(S): Oncorhynchus mykiss

SUBMITTER: Makoto Hayashi

PROVIDER: E-GEOD-38647 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Mammalian spermatogenesis is a complex biological process that occurs within a highly organized tissue, the seminiferous epithelium. The coordinated maturation of spermatogonia, spermatocytes and spermatids suggests the existence of precise programs of gene expression in these cells as well as in their neighboring somatic Sertoli cells. The objective of this study was to elucidate genes encoding the proteins that execute these programs. Rat seminiferous tubules at stages I, II-III, IV-V, VI, VIIa,b, VIIc,d, VIII, IX-XI, XII, XIII-XIV of the cycle were isolated by microdissection and Sertoli cells, spermatogonia plus early spermatocytes, pachytene spermatocytes and spermatids were purified from enzymatically-dispersed testes. Microarray analysis using Rat Genome 230 2.0 arrays identified a total of 16,971 probe sets that recognized transcripts. A comparison with the transcriptome of other tissues identified 398 testis-specific probe sets, which therefore are potential targets for the development of new contraceptives. Sequential waves of cell and stage-specific gene expression are associated with progression of germ cells through the stages of the cycle of the seminiferous epithelium and 1612 probe sets recognized transcripts whose expressions varied at least 4-fold across the stages of the cycle. Pathway analyses reveal that entire biological processes are regulated cyclically in testicular cells. Important among these are cell cycle and DNA repair. Thus, stage-specific gene expression is a widespread and fundamental characteristic of spermatogenic cells and Sertoli cells. Experiment Overall Design: Seminiferous tubules at the following stages or groups of stages were isolated by transillumination-assisted microdissection: I, II-III, IV-V, VI, VIIa,b, VIIc,d, VIII, IX-XI, XII, XIII-XIV. Twenty-five segments of tubules, 2 m in length, were collected at each stage or group of stages from each rat. Tubules from two separate animals were pooled to form one sample from each stage or groups of stages. A total of 5 independent samples per stage or groups of stages were analyzed. Mature Sertoli cells were isolated to about 85% purity (n=4). Three groups of germ cells were isolated by centrifugal elutriation from enzyme-dispersed testes: round spermatids (n=4), pachytene spermatocytes (n=4) and spermatogonia plus early spermatocytes (n=3) . Purity of these cells has been shown to vary between 85% to 95%

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Identification of type A spermatogonia specific transcriptome in rainbow trout immature testis

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